title: The monovalent cation leak in overhydrated stomatocytic red blood cells results from amino acid substitutions in the Rh-associated glycoprotein.
abstract: Overhydrated hereditary stomatocytosis (OHSt) is a rare dominantly inherited hemolytic anemia characterized by a profuse membrane leak to monovalent cations. Here, we show that OHSt red cell membranes contain slightly reduced amounts of Rh-associated glycoprotein (RhAG), a putative gas channel protein. DNA analysis revealed that the OHSt patients have 1 of 2 heterozygous mutations (t182g, t194c) in RHAG that lead to substitutions of 2 highly conserved amino acids (Ile61Arg, Phe65Ser). Unexpectedly, expression of wild-type RhAG in Xenopus laevis oocytes induced a monovalent cation leak; expression of the mutant RhAG proteins induced a leak about 6 times greater than that in wild type. RhAG belongs to the ammonium transporter family of proteins that form pore-like structures. We have modeled RhAG on the homologous Nitrosomonas europaea Rh50 protein and shown that these mutations are likely to lead to an opening of the pore. Although the function of RhAG remains controversial, this first report of functional RhAG mutations supports a role for RhAG as a cation pore.
authors.affiliation: Bristol Institute for Transfusion Sciences, National Health Service Blood and Transplant, Filton, Bristol, United Kingdom. lesley.bruce@nhsbt.nhs.uk
authors.forename: Lesley J
authors.initials: LJ
authors.lastname: Bruce
authors.forename: Hélène
authors.initials: H
authors.lastname: Guizouarn
authors.forename: Nicholas M
authors.initials: NM
authors.lastname: Burton
authors.forename: Nicole
authors.initials: N
authors.lastname: Gabillat
authors.forename: Joyce
authors.initials: J
authors.lastname: Poole
authors.forename: Joanna F
authors.initials: JF
authors.lastname: Flatt
authors.forename: R Leo
authors.initials: RL
authors.lastname: Brady
authors.forename: Franck
authors.initials: F
authors.lastname: Borgese
authors.forename: Jean
authors.initials: J
authors.lastname: Delaunay
authors.forename: Gordon W
authors.initials: GW
authors.lastname: Stewart
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000743:Anemia, Hemolytic / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D000818:Animals; D001798:Blood Proteins / Q000235:genetics* / Q000378:metabolism; D002414:Cations, Monovalent / Q000378:metabolism*; D004910:Erythrocyte Membrane / Q000378:metabolism; D004912:Erythrocytes / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D015151:Immunoblotting; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D043370:Nitrosomonas europaea / Q000378:metabolism; D009865:Oocytes / Q000166:cytology / Q000378:metabolism; D011487:Protein Conformation; D012204:Rh-Hr Blood-Group System / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D014982:Xenopus laevis / Q000378:metabolism
chemical_list: D001798:Blood Proteins; D002414:Cations, Monovalent; D008562:Membrane Glycoproteins; C498523:RHAG protein, human; D012204:Rh-Hr Blood-Group System